Francisella tularensis is the causative agent of respiratory tularemia, a debilitating disease of humans. This bacterial pathogen has been listed as Category A Select Agent owing to its extreme virulence and the ease of its dissemination via aerosol route. To date there is no effective immune therapy or vaccine licensed for prevention of this disease. Although clinical and experimental studies have shown that Th-1 type of host immune responses are protective, bacterial antigens driving these responses are not well defined. Identification of such antigens will aide in formulating effective prevention strategies fr this debilitating disease. This encompasses the goal of the proposed studies. For this, we aim to utilize a novel approach of comparing the immunodominant protein profile of Francisella using sera from mice inoculated with a mutant Francisella strain that is attenuated for causing the infection but does not protect the mice from a lethal challenge with the virulent wild-type organisms and sera from mice inoculated with mutants that not only are attenuated but also protect the mice from lethal challenge. Based on our preliminary studies we believe that this unique approach will identify Francisella proteins associated only with the protective response which can then be utilized as vaccine candidates. In this line, we are armed with a collection of attenuated/non-protective and attenuated/protective mutants of Francisella which will be used to inoculate the mice and the sera collected from these mice will be used to probe total proteins of Francisella followed by sequencing and identification of immunodominant proteins reactive only to the sera from mice inoculated with protective mutants (Aim 1). These proteins will then be produced as recombinant fusion proteins and tested for their protective efficacy against pulmonary infection with virulent Francisella strains (Aim 2). We believe that these studies will uncover Francisella proteins capable of generating protective anti-Francisella immunity thus serving as candidates for a subunit vaccine against this pathogen. Additionally, this novel strategy of comparative immunoproteomics may serve as a platform to identify vaccine candidates for other bacterial pathogens as well. The outcome of proposed studies is expected to take the Francisella subunit vaccine research a step further.